Abstract/Project Summary During aging, the ability of neural stem cells (NSCs) in the brain to form new neurons is reduced, but the molecular mechanisms underlying the deterioration of NSC function remain unclear. There is currently a critical need to understand the mechanisms by which NSCs are activated to form neurons, and why this process declines with age. The long term goal is to identify the mechanisms responsible for the loss of NSC function with age, and discover interventions that harness the regenerative capacity of these cells to increase cognitive function in aged and diseased individuals. The objective of this proposal is to identify the mechanisms by which the pro-longevity transcription factor FOXO3 maintains NSCs during aging. The central hypothesis is that FOXO3 directly regulates a network of target genes and pathways that are critical for preserving NSCs during aging. This hypothesis will be tested by pursuing the following specific aims: 1) Determine the mechanisms underlying FOXO3-mediated NSC quiescence. 2) Determine how changes in the FOXO3 network underlie the decline in NSC function with age. 3) Determine the extent to which FOXO3 can preserve stem cells in vivo during aging. The first aim will be accomplished by combining a model of primary adult mouse NSC quiescence with loss of function and overexpression approaches to test the hypothesis that FOXO3 directly promotes quiescence by regulating specific genes and pathways. The second aim will be performed using methods to test the extent to which levels, activity, or binding sites downstream of FOXO3 are responsible for observed gene expression changes in aging NSCs. The third aim will be accomplished using a transgenic mouse overexpressing FOXO3 under endogenous regulation to test the hypothesis that increasing levels of FOXO3 maintains NSCs in a quiescent state during aging, thereby increasing the pool of NSCs in old mice. The outcome of this project will be the identification of the mechanisms by which FOXO3 regulates NSC function, how these mechanisms deteriorate with age, and reveal a strategy that reverses the loss of NSCs in aging mice. This work is significant because it will determine why NSC activation is reduced in the aged brain, and reveal strategies to reverse it. This proposed research is innovative because it will be the first to elucidate a direct transcriptional mechanism to promote adult NSC quiescence. This work will provide key mechanistic insight into how gene networks are coordinated in young and aging NSCs, and have the potential to reveal new mechanisms underlying cognitive decline during aging.